Analysis of Testosterone Using the Xevo TQ-XS for Clinical Research

Importance of the Topic

Serum testosterone measurement is critical in clinical research for assessing endocrine and metabolic disorders. Conventional immunoassays often suffer from cross-reactivity with structurally related steroids, leading to poor accuracy and reproducibility at low concentrations. High-sensitivity LC-MS/MS methods overcome these limitations by combining selective sample preparation, chromatographic separation, and mass-spectrometric detection.

Objectives and Study Overview

The primary aim of this study was to demonstrate the analytical sensitivity, precision, and quantitative performance of the Waters Xevo TQ-XS tandem quadrupole mass spectrometer for testosterone determination in human serum using only 100 µL sample volume. Key performance indicators included lower limit of quantification, linear dynamic range, method precision, and bias against certified reference materials.

Methodology

Serum samples (100 µL) were protein-precipitated with methanol, diluted with water, and processed on an Oasis PRiME HLB µElution Plate. After two wash steps (0.1% ammonia in 35% methanol; 0.1% formic acid in 35% methanol), analytes were eluted in 85:15 acetonitrile–methanol, diluted with water, and directly injected.

Instrumentation Used

• Waters ACQUITY UPLC I-Class System
• ACQUITY UPLC HSS T3 Column with VanGuard T3 pre-column
• Waters Xevo TQ-XS Tandem Quadrupole Mass Spectrometer
• Oasis PRiME HLB µElution Plate

Main Results and Discussion

– Lower limit of quantification: 5 pg/mL on-column (0.017 nmol/L) with RSD <3% for peak area.
– Calibration linearity from 0.005–20 ng/mL (r2 >0.997).
– Total precision (n=18) ≤4.4% across 0.02–14 ng/mL.
– Method bias within ±3.3% against ERM DA345a and DA346a reference materials.
– Signal-to-noise ratios exceeded 160 at 5 pg/mL spikes and remained robust across the calibration range.

Benefits and Practical Applications

– Ultra-sensitive detection of testosterone in low-volume serum critical for pediatric, anti-doping, and endocrinology studies.
– High precision and wide dynamic range enable reliable quantification across physiologically relevant concentrations.
– Streamlined sample preparation and µElution reduce solvent use and processing time.

Future Trends and Opportunities

Advances in ion transfer optics and detector technology will continue to push detection limits lower. Integration of automated on-line sample cleanup, coupling with high-resolution MS, and expansion to multiplexed steroid panels will broaden clinical research utility. Miniaturized workflows may enable bedside or point-of-care endocrine monitoring.

Conclusion

The Xevo TQ-XS platform delivers exceptional sensitivity, precision, and accuracy for serum testosterone analysis with minimal sample volume. This robust LC-MS/MS approach addresses limitations of immunoassays and supports rigorous quantitative studies in clinical and research settings.

References

No external literature references were provided in the original document.